Carotenoid determination in biological microsamples using liquid chromatography with a coulometric electrochemical...

Numerous epidemiological studies have linked carotenoids to cancer preventive processes, thereby increasing interest in levels of these micronutrients in human tissue and serum. Conventional analyses of these biological tissues employ liquid chromatography (LC) with ultraviolet and visible absorbance (UV-VIS) detection. However, this type of carotenoid analysis does not provide adequate sensitivity for very small sample sizes, such as microscale biopsies, when only small quantities of tissue are available. Electrochemical detection (ECD) is a useful alternative to conventional UV-VIS detection methods for LC analysis of carotenoids in cases where high sensitivity is necessary. Both hydrocarbon (beta-carotene and alpha-carotene) and oxygenated carotenoids (lutein, zeaxanthin, and beta-cryptoxanthin) were detected at electrical potential settings between 220 and 520 mV. The generated electrochemical array data (hydrodynamic voltammograms) can be used to identify carotenoids as well as to differentiate between trans and cis configurations. Detection limits for beta-carotene by ECD were measured at 10 fmol representing approximately a 100- to 1000-fold increase over conventional LC-UV-VIS techniques. The developed methodology was applied successfully to microscale analysis of biological tissues.